What you need to know about genetics All organisms have DNA and RNA as genetic material All organisms use the same nucl

1. What you need to know about genetics All organisms have DNA and RNA as genetic material All organisms use the same nucleotides All organisms replicate, transcribe and translate DNA

2. The central dogma of molecular biology

3. What is different in prokaryotes from eukaryotes? The prokaryotic genome is circular, so it is repli- cated differently than linear eukaryotic DNA What is gene expression? transcription: synthesis of RNA from a DNA template translation: formation of a protein (amino acid sequence) from RNA sequence How does this process differ in prokaryotes?

4. In prokaryotes, an mRNA molecule can carry information for several genes e.g., all of the genes in a biosynthetic pathway (this doesn’t happen in eukaryotes) The RNA polymerase in constructed differently in prokaryotes Process consists of initiation, elongation, termination Same steps are seen in translation

5. What are the differences? In eukaryotes, RNA is modified and transported from the nucleus The ribosomes are different (70S in prokaryotes, 80S in eukaryotes The proteins and enzymes are different that make transcription and translation occur

6. The entire genomes (DNA sequences) of many bacteria are known >1000 times smaller than human genome

7. Gene expression is carefully controlled. Bacteria must be able to adapt to environment and divide quickly Some genes are constitutive (always expressed) Some are inducible (only expressed when needed) example: enzymes that metabolize lactose Some are repressible ( expressed most of the time, but can be turned off) example: enzymes involved in biosynthesis These genes are organized into operons

8. Operons: a group of genes expressed together

9. Repressor: binds to operator (a region of DNA adjacent to genes) If something is bound to operator, trasncription cannot occur Inducer: binds to repressor so it can’t bind to operator Therefore transcription can take place Example: lactose operon. genes are expressed only if lactose (inducer) is present

10. Biosynthetic genes are also arranged in operons In this case, the repressor is the end product of the pathway

11. Some fundamentals of bacterial genetics Bacterial DNA (like any DNA) can be altered by mutations Mutations can result in changes in proteins diversity acquisition of resistance new traits can be transmitted to other microbes

12. What happens when DNA is mutated? Bases (nucleotides) are changed (base substitution) Bases are inserted or deleted Sequences of DNA move around the genome (transposons)

13. Point mutation: a single amino acid is changed

14. How many amino acids are changed?

15. Mutations can occur spontaneously, or can be induced By chemicals By radiation

16. DNA damage occurs fairly frequently Cells have several repair pathways available to them Repair of errors in base incorporation Repair of thymine dimers (due to UV irradiation) Repair of damaged DNA Bypass of damaged DNA (SOS repair)

17. In bacteria, mutants can be identified by plating on special minimal medium Auxotrophs have lost the ability to synthesize certain substances (Prototrophs: retain the ability to do so) A variety of selective strategies have been developed

19. The Ames test uses bacteria to test chemicals for capacity to cause mutations, as well as carcinogens (cancer-causing chemicals) Much more rapid and inexpensive than testing animals Results have been reliable and reproducible Several tester strains are used, since chemicals can cause point or frameshift mutations (but not both)

20. p. 202

21. How does gene transfer occur in bacteria? Only part of the bacterial DNA is transferred The new DNA replaces existing DNA (homologous recombination) Transformation- uptake of “naked” DNA Transduction- by bacteriophages Conjugation- bacterial cells come in direct contact with each other. Plasmid is often transferred

23. Transformant identified by selection

24. Recipient bacteria must be “competent” to take up and incorporate DNA Few strains of bacteria are naturally competent (generally Gram positive) Bacteria can be made artificially competent calcium solutions electric current

25. Transduction: bacteriophage transfer DNA from one bacterium to another

27. Conjugation Direct contact between donor and recipient must occur Sex pilus is encoded by fertility (F) plasmid

29. Note role of plasmids in conjugation What is a plasmid? Small, circular double-stranded DNA Common in prokaryotes May vary widely in size Can encode a variety of genes; usually not essential bacterial genes but may give bacterium new properties

31. R plasmids- resistance to antibiotics, metals Virulence factors (that make bacteria pathogenic, or disease causing) Transposons can insert themselves into genome (and out of it) Thus bacteria have many ways of obtaining new genes to enhance survival

32. Natural methods of gene transfer have been modified in biotechnology Deliberate introduction of genes into recipient bacteria or other cells Genes could not otherwise be exchanged “recombinant DNA”

34. Plasmid may be able to replicate frequently, so many copies of the DNA are made The gene may be expressed, so the protein product in made in large quantities e.g., recombinant human insulin, as opposed to extracts from pig pancreases Genetic analysis determining DNA sequences analyzing gene products expressing DNA in a variety of cells What do the proteins do?

35. Summary DNA replication, transcription and translation occur in all cells Prokaryotes have a single, circular DNA genome They have different enzymes and ribosomes but use the same nucleotides They acquire new traits through incorporating mutations. These can be analyzed through selection techniques

36. Bacteria do not reproduce sexually but can acquire new DNA through transformation, transduction or conjugation These natural processes have been modified so that DNA can be deliberately incorpor- ated into host microbes- even genes that would normally never be transferred this way